When proteins useful as pharmaceuticals are produced with the recombinant DNA technique, use of animal cells enables complicated post-translational modification and folding which prokaryotic cells can not perform. Therefore, animal cells are frequently used as host cells for producing recombinant proteins.
Recently, a large number of biopharmaceuticals, such as antibodies and physiologically active proteins, have been developed. Techniques that permit efficient production of recombinant proteins by animal cells lead to cost reduction of biopharmaceuticals and promise their stable supply to patients.
Under these circumstances, a method of protein production with higher production efficiency is desired.
Alanine is one of the proteinogenic amino acids, and is a non-essential amino acid. In a living body, it is biosynthesized by transfer of an amino group from glutamate to pyruvate, and is degraded by a reverse reaction.
As an alanine degrading enzyme, alanine aminotransferase (EC 2.6.1.2.) (Non-patent document 1) has been known. This enzyme transfers an amino group from alanine to 2-oxoglutarate to synthesize glutamate. Alanine aminotransferase is also called glutamic-pyruvic transaminase, which is abbreviated as GPT (Non-patent document 2). GPT and GOP (aspartate aminotransferase) are enzymes found in the liver. Since GPT and GOP are released into the blood when hepatic cells are destroyed, the liver is diagnosed to have some kind of disorder when abnormally high levels of GPT and GOT are observed.
As shown above, alanine aminotransferase is used as a marker of hepatic function. However, it has not been known how host cells such as CHO cells behave if alanine aminotransferase is strongly expressed in them.
[Non-Patent Document 1]
Sanjay B. J., et. al., Hepatology (2004) 39(5), 1297-1302
[Non-Patent Document 2]
Melanie M. S., et. al., Genomics (1997) 40, 247-252